The development of autoimmunity involves the failure of the mechanisms that regulate the ability to discriminate self from non-self. The primary means of regulating self-tolerance is through the deletion of self-reactive cells in the thymus. However, this mechanism is not perfect and auto-reactive clones do escape into the periphery. Peripheral tolerance is generated through a variety of mechanisms including T cell anergy and T cell indifference or ignorance. Recently, another, more active, mechanism of tolerance induction has been identified that is controlled by a population of regulatory T cells that actively suppress the function of auto-reactive T cells. These T cells, know as TR cells, have a characteristic cell-surface phenotype and fail to respond to typical T cell stimulation regimens in vitro. However, they have the ability to inhibit the development of autoimmunity when transferred into the appropriate host. However, the mechanism by which these cells perform their tasks is as yet unclear. Recent work has shown that the forkhead/winged-helix protein FoxP3 is expressed predominantly in TR cells, and is both necessary and sufficient for their development and function. The finding that FoxP3 expression determines TR cell fate, as well as the ability of ectopic FoxP3 expression to convert naive T cells into TR cells, suggests that FoxP3 may be useful in cell-based therapies for autoimmune diseases. As a first step, the experiments in this proposal are designed to test the feasibility of using ectopic FoxP3 expression as a means of converting both normal and pathogenic T cells to Tg-like cells. We will use an in vitro cell culture system to determine whether introduction of FoxP3 into primary antigen-specific human T cells can convert these cells into antigen-specific TR cells. We will also use two adoptive transfer models of type I diabetes in the mouse to test the ability of ectopic FoxP3 expression to convert diabetogenic T cells into TR cells. These experiments will provide valuable information on the efficacy of FoxP3 as a tool for therapeutic intervention in autoimmune diabetes.